The Salt-Inducible Kinases: Emerging Metabolic Regulators

Salt-inducible kinases (SIKs) are related to AMP-activated protein kinase (AMPK), but do not appear to be involved in the sensing and regulation of cellular energy levels. SIKs and other AMPK-related kinases all require phosphorylation of their kinase domain T-loop to be active. Unlike AMPK however, SIKs are constitutively T-loop phosphorylated and their intrinsic kinase activity is thus rarely regulated in response to extracellular signals. SIKs are however phosphorylated, for example, by PKA, outside the kinase domain, leading to changes in their cellular function. SIK activity is strictly required for the inhibition of glucose production in mouse liver; however, the contribution of different SIK isoforms to this function in vivo is not yet clear. SIK2 is downregulated in adipose tissue from obese and insulin-resistant individuals and might contribute to reduced insulin sensitivity and glucose uptake in adipocytes during obesity. The discovery of liver kinase B1 (LKB1) as an upstream kinase for AMP-activated protein kinase (AMPK) led to the identification of several related kinases that also rely on LKB1 for their catalytic activity. Among these, the salt-inducible kinases (SIKs) have emerged as key regulators of metabolism. Unlike AMPK, SIKs do not respond to nucleotides, but their function is regulated by extracellular signals, such as hormones, through complex LKB1-independent mechanisms. While AMPK acts on multiple targets, including metabolic enzymes, to maintain cellular ATP levels, SIKs primarily regulate gene expression, by acting on transcriptional regulators, such as the cAMP response element-binding protein-regulated transcription coactivators and class IIa histone deacetylases. This review describes the development of research on SIKs, from their discovery to the most recent findings on metabolic regulation. The discovery of liver kinase B1 (LKB1) as an upstream kinase for AMP-activated protein kinase (AMPK) led to the identification of several related kinases that also rely on LKB1 for their catalytic activity. Among these, the salt-inducible kinases (SIKs) have emerged as key regulators of metabolism. Unlike AMPK, SIKs do not respond to nucleotides, but their function is regulated by extracellular signals, such as hormones, through complex LKB1-independent mechanisms. While AMPK acts on multiple targets, including metabolic enzymes, to maintain cellular ATP levels, SIKs primarily regulate gene expression, by acting on transcriptional regulators, such as the cAMP response element-binding protein-regulated transcription coactivators and class IIa histone deacetylases. This review describes the development of research on SIKs, from their discovery to the most recent findings on metabolic regulation. AMPK (see Glossary) is an evolutionary conserved sensor and regulator of cellular energy levels that coordinates metabolic pathways to balance ATP consumption with production. Identification of LKB1, a tumor suppressor, as a major upstream regulator for AMPK provided an unanticipated novel link between cancer and metabolism. Furthermore, it uncovered the existence of another 12 kinases, closely related to AMPK, that are also activated by LKB1 and have a similar substrate specificity in vitro [1Lizcano J.M. et al.LKB1 is a master kinase that activates 13 kinases of the AMPK subfamily, including MARK/PAR-1.EMBO J. 2004; 23: 833-843Crossref PubMed Scopus (1058) Google Scholar]. At the time of this discovery, the expression, regulation, and functions of many AMPK-related kinases were still poorly characterized [2Alessi D.R. et al.LKB1-dependent signaling pathways.Annu. Rev. Biochem. 2006; 75: 137-163Crossref PubMed Scopus (636) Google Scholar]. However, it soon became clear that the LKB1 signaling network plays a role in the control of diverse biological processes, beyond nutrient sensing/metabolism and tumor suppression [3Bright N.J. et al.The regulation and function of mammalian AMPK-related kinases.Acta Physiol. (Oxf). 2009; 196: 15-26Crossref PubMed Scopus (126) Google Scholar]. It was also apparent that AMPK-related kinases are distinct from AMPK in that they are not primarily activated by cellular energy levels, but are subject to complex regulation by multiple signals and kinases [3Bright N.J. et al.The regulation and function of mammalian AMPK-related kinases.Acta Physiol. (Oxf). 2009; 196: 15-26Crossref PubMed Scopus (126) Google Scholar]. The mammalian SIK family of AMPK-related kinases consists of three isoforms, SIK1, SIK2, and SIK3, that were identified and cloned just before the publication by Lizcano et al. in 2004 (Box 1) [4Wang Z. et al.Cloning of a novel kinase (SIK) of the SNF1/AMPK family from high salt diet-treated rat adrenal.FEBS Lett. 1999; 453: 135-139Crossref PubMed Scopus (118) Google Scholar, 5Feldman J.D. et al.The salt-inducible kinase, SIK, is induced by depolarization in brain.J. Neurochem. 2000; 74: 2227-2238Crossref PubMed Scopus (47) Google Scholar, 6Xia Y. et al.The new serine-threonine kinase, Qik, is a target of the Qin oncogene.Biochem. Biophys. Res. Commun. 2000; 276: 564-570Crossref PubMed Scopus (10) Google Scholar, 7Ruiz J.C. et al.Identification of novel protein kinases expressed in the myocardium of the developing mouse heart.Mech. Dev. 1994; 48: 153-164Crossref PubMed Scopus (48) Google Scholar, 8Horike N. et al.Adipose-specific expression, phosphorylation of Ser794 in insulin receptor substrate-1, and activation in diabetic animals of salt-inducible kinase-2.J. Biol. Chem. 2003; 278: 18440-18447Crossref PubMed Scopus (118) Google Scholar, 9Screaton R.A. et al.The CREB coactivator TORC2 functions as a calcium- and cAMP-sensitive coincidence detector.Cell. 2004; 119: 61-74Abstract Full Text Full Text PDF PubMed Scopus (511) Google Scholar]. An important early discovery was that SIKs inhibit gene expression controlled by the master transcription factor cAMP response element-binding protein (CREB) [10Takemori H. et al.ACTH-induced nucleocytoplasmic translocation of salt-inducible kinase. Implication in the protein kinase A-activated gene transcription in mouse adrenocortical tumor cells.J .Biol. Chem. 2002; 277: 42334-42343Crossref PubMed Scopus (63) Google Scholar, 11Doi J. et al.Salt-inducible kinase represses cAMP-dependent protein kinase-mediated activation of human cholesterol side chain cleavage cytochrome P450 promoter through the CREB basic leucine zipper domain.J. Biol. Chem. 2002; 277: 15629-15637Crossref PubMed Scopus (47) Google Scholar]. This provided the first clue to a metabolic role for SIK isoforms. The subsequent identification of the CREB-regulated transcription coactivators (CRTCs) (reviewed in [12Altarejos J.Y. Montminy M. CREB and the CRTC co-activators: sensors for hormonal and metabolic signals.Nat. Rev. Mol. Cell Biol. 2011; 12: 141-151Crossref PubMed Scopus (716) Google Scholar]) as substrates for SIKs further triggered research in this area. By now, SIKs have emerged as regulators of various metabolic pathways that maintain glucose- and lipid homeostasis. They also respond to metabolic hormones (Box 2) and thus play a role in hormonal control of metabolism. Moreover, additional substrates, in particular the class IIa histone deacetylases (HDACs), have been identified that likely mediate metabolic effects of SIKs.Box 1Discovery of SIKsDuring 1999–2000, Sik was identified as a gene that is rapidly induced in response to various stimuli in different cell types. Upon identification of additional isoforms, this gene and its gene product were later denoted Sik1. For clarification, the first SIK isoform to be identified is termed SIK1 throughout this review article. Wang et al. [4Wang Z. et al.Cloning of a novel kinase (SIK) of the SNF1/AMPK family from high salt diet-treated rat adrenal.FEBS Lett. 1999; 453: 135-139Crossref PubMed Scopus (118) Google Scholar] sought to identify factors that regulate function of the adrenal cortex in response to changes in plasma Na+/K+ balance and screened genes that were induced in rat adrenal gland following a Na+- or K+-enriched diet. A novel cDNA clone, termed salt-inducible kinase (Sik), encoding a 776-amino acid polypeptide with substantial similarity to the AMPK/sucrose non-fermenting kinase (SNF1, the yeast homolog of mammalian AMPK) family, emerged from the screen. They also found that Sik1 mRNA expression is rapidly stimulated by adrenocorticotropic hormone in mouse and rat adrenal cells. Feldman et al. [5Feldman J.D. et al.The salt-inducible kinase, SIK, is induced by depolarization in brain.J. Neurochem. 2000; 74: 2227-2238Crossref PubMed Scopus (47) Google Scholar] were searching for genes in neuronal cells that were preferentially induced by membrane depolarization. They identified a gene product that is induced by potassium, forskolin, or a calcium ionophore and hence termed it KID-2 (kinase-induced by depolarization-2). Xia et al. [6Xia Y. et al.The new serine-threonine kinase, Qik, is a target of the Qin oncogene.Biochem. Biophys. Res. Commun. 2000; 276: 564-570Crossref PubMed Scopus (10) Google Scholar] were studying the role of Qin, a gene shown to induce oncogenic transformation, and sought to identify genes that were differentially expressed in chicken embryo fibroblasts transformed by Qin. One of the up-regulated Qin targets identified was a protein kinase termed QIK (Qin-induced kinase) that displayed high homology to SIK1 cloned by Wang et al. [4Wang Z. et al.Cloning of a novel kinase (SIK) of the SNF1/AMPK family from high salt diet-treated rat adrenal.FEBS Lett. 1999; 453: 135-139Crossref PubMed Scopus (118) Google Scholar] just before their publication. It should also be noted that in 1994, Ruiz and colleagues [7Ruiz J.C. et al.Identification of novel protein kinases expressed in the myocardium of the developing mouse heart.Mech. Dev. 1994; 48: 153-164Crossref PubMed Scopus (48) Google Scholar] had also identified a putative serine/threonine kinase most similar to SNF1, named myocardial SNF-like kinase (msk).Identification of Sik1 prompted Okamoto and colleagues to search for its closely related genes. Analysis of human and mouse genomic databases led them to identify such genes, which they termed Sik2 and Sik3 [8Horike N. et al.Adipose-specific expression, phosphorylation of Ser794 in insulin receptor substrate-1, and activation in diabetic animals of salt-inducible kinase-2.J. Biol. Chem. 2003; 278: 18440-18447Crossref PubMed Scopus (118) Google Scholar, 9Screaton R.A. et al.The CREB coactivator TORC2 functions as a calcium- and cAMP-sensitive coincidence detector.Cell. 2004; 119: 61-74Abstract Full Text Full Text PDF PubMed Scopus (511) Google Scholar]. The Sik1 gene is located on human chromosome 21, whereas both Sik2 and Sik3 genes are localized on chromosome 11. SIKs are evolutionary conserved; Drosophila melanogaster and Caenorhabditis elegans both have SIK homologs, termed dSIK and Kin-29, respectively [13Lanjuin A. Sengupta P. Regulation of chemosensory receptor expression and sensory signaling by the KIN-29 Ser/Thr kinase.Neuron. 2002; 33: 369-381Abstract Full Text Full Text PDF PubMed Scopus (48) Google Scholar, 14Okamoto M. et al.Salt-inducible kinase in steroidogenesis and adipogenesis.Trends Endocrinol. Metab. 2004; 15: 21-26Abstract Full Text Full Text PDF PubMed Scopus (72) Google Scholar, 15Wang B. et al.The insulin-regulated CREB coactivator TORC promotes stress resistance in Drosophila.Cell Metab. 2008; 7: 434-444Abstract Full Text Full Text PDF PubMed Scopus (75) Google Scholar, 16Choi S. et al.Drosophila salt-inducible kinase (SIK) regulates starvation resistance through cAMP-response element-binding protein (CREB)-regulated transcription coactivator (CRTC).J. Biol. Chem. 2011; 286: 2658-2664Crossref PubMed Scopus (39) Google Scholar].Box 2Structure and Regulation of SIKs by PhosphorylationAll three SIK isoforms share a highly homologous N-terminal kinase domain, but C-terminal regions are less conserved and differ in length (Figure I). SIK isoforms also possess a ubiquitin-associated domain (Figure I) that appears to play a role in allowing SIKs (and other AMPK-related kinases) to be activated by LKB1 [17Jaleel M. et al.Identification of the sucrose non-fermenting related kinase SNRK, as a novel LKB1 substrate.FEBS Lett. 2005; 579: 1417-1423Crossref PubMed Scopus (116) Google Scholar, 18Rider M.H. The ubiquitin-associated domain of AMPK-related protein kinases allows LKB1-induced phosphorylation and activation.Biochem. J. 2006; 394: e7-e9Crossref PubMed Scopus (6) Google Scholar].Phosphorylation of the Kinase Domain by LKB1Like other AMPK-related kinases, SIKs are phosphorylated by LKB1 on a highly conserved threonine residue (homologous to Thr172 in AMPK) in their activation loop [1Lizcano J.M. et al.LKB1 is a master kinase that activates 13 kinases of the AMPK subfamily, including MARK/PAR-1.EMBO J. 2004; 23: 833-843Crossref PubMed Scopus (1058) Google Scholar, 17Jaleel M. et al.Identification of the sucrose non-fermenting related kinase SNRK, as a novel LKB1 substrate.FEBS Lett. 2005; 579: 1417-1423Crossref PubMed Scopus (116) Google Scholar]. This phosphorylation is essential for their catalytic activity, but not known to be affected in response to external stimuli or by low cellular energy conditions [1Lizcano J.M. et al.LKB1 is a master kinase that activates 13 kinases of the AMPK subfamily, including MARK/PAR-1.EMBO J. 2004; 23: 833-843Crossref PubMed Scopus (1058) Google Scholar, 19Sakamoto K. et al.Activity of LKB1 and AMPK-related kinases in skeletal muscle: effects of contraction, phenformin, and AICAR.Am. J. Physiol. Endocrinol. Metab. 2004; 287: E310-E317Crossref PubMed Scopus (264) Google Scholar] like AMPK. The most widely received model is that LKB1 is constitutively active; therefore, its dependent kinases (except AMPK) are also constitutively phosphorylated (on the activation loop) and active in cells.Phosphorylation Outside of the Kinase Domain in Response to Metabolic CuesMultiple studies have shown that SIKs are phosphorylated at a C-terminal serine residue (SIK1, Ser577; SIK2, Ser587; SIK3, Ser551) in response to agents that elevate intracellular cAMP levels and thus activate protein kinase A (PKA) [10Takemori H. et al.ACTH-induced nucleocytoplasmic translocation of salt-inducible kinase. Implication in the protein kinase A-activated gene transcription in mouse adrenocortical tumor cells.J .Biol. Chem. 2002; 277: 42334-42343Crossref PubMed Scopus (63) Google Scholar, 20Henriksson E. et al.The AMPK-related kinase SIK2 is regulated by cAMP via phosphorylation at Ser358 in adipocytes.Biochem. J. 2012; 444: 503-514Crossref PubMed Scopus (49) Google Scholar, 21Patel K. et al.The LKB1-salt-inducible kinase pathway functions as a key gluconeogenic suppressor in the liver.Nat. Commun. 2014; 5: 4535-4550Crossref PubMed Scopus (104) Google Scholar, 22Sonntag T. et al.14-3-3 proteins mediate inhibitory effects of cAMP on salt-inducible kinases (SIKs).FEBS J. 2018; 285: 467-480Crossref PubMed Scopus (30) Google Scholar, 23Berggreen C. et al.cAMP-elevation mediated by beta-adrenergic stimulation inhibits salt-inducible kinase (SIK) 3 activity in adipocytes.Cell Signal. 2012; 24: 1863-1871Crossref PubMed Scopus (23) Google Scholar]. Detailed studies of glucagon- and catecholamine-induced phosphorylation of SIK2 have been performed in hepatocytes and adipocytes, respectively, and these studies report additional PKA phosphorylation sites (Ser343, Ser358, Thr484, and Ser587) [20Henriksson E. et al.The AMPK-related kinase SIK2 is regulated by cAMP via phosphorylation at Ser358 in adipocytes.Biochem. J. 2012; 444: 503-514Crossref PubMed Scopus (49) Google Scholar, 21Patel K. et al.The LKB1-salt-inducible kinase pathway functions as a key gluconeogenic suppressor in the liver.Nat. Commun. 2014; 5: 4535-4550Crossref PubMed Scopus (104) Google Scholar]. A large body of evidence indicates that cAMP-induced phosphorylation serves to restrict SIK cellular activity [9Screaton R.A. et al.The CREB coactivator TORC2 functions as a calcium- and cAMP-sensitive coincidence detector.Cell. 2004; 119: 61-74Abstract Full Text Full Text PDF PubMed Scopus (511) Google Scholar, 10Takemori H. et al.ACTH-induced nucleocytoplasmic translocation of salt-inducible kinase. Implication in the protein kinase A-activated gene transcription in mouse adrenocortical tumor cells.J .Biol. Chem. 2002; 277: 42334-42343Crossref PubMed Scopus (63) Google Scholar, 24Dentin R. et al.Insulin modulates gluconeogenesis by inhibition of the coactivator TORC2.Nature. 2007; 449: 366-369Crossref PubMed Scopus (324) Google Scholar, 25Muraoka M. et al.Involvement of SIK2/TORC2 signaling cascade in the regulation of insulin-induced PGC-1alpha and UCP-1 gene expression in brown adipocytes.Am. J. Physiol. Endocrinol. Metab. 2009; 296: E1430-E1439Crossref PubMed Scopus (41) Google Scholar], however with mechanisms that differ between SIK isoforms and that in several aspects remain unclear. Rather than a change in intrinsic activity, altered substrate availability is likely involved. Indeed, SIK1 undergoes phosphorylation-dependent nucleocytoplasmic shuttling [10Takemori H. et al.ACTH-induced nucleocytoplasmic translocation of salt-inducible kinase. Implication in the protein kinase A-activated gene transcription in mouse adrenocortical tumor cells.J .Biol. Chem. 2002; 277: 42334-42343Crossref PubMed Scopus (63) Google Scholar]. Moreover, the phosphorylation of SIK2 and SIK3 was shown to induce their binding to 14-3-3 proteins, which coincides with reduced substrate binding [20Henriksson E. et al.The AMPK-related kinase SIK2 is regulated by cAMP via phosphorylation at Ser358 in adipocytes.Biochem. J. 2012; 444: 503-514Crossref PubMed Scopus (49) Google Scholar, 22Sonntag T. et al.14-3-3 proteins mediate inhibitory effects of cAMP on salt-inducible kinases (SIKs).FEBS J. 2018; 285: 467-480Crossref PubMed Scopus (30) Google Scholar, 26Henriksson E. et al.SIK2 regulates CRTCs, HDAC4 and glucose uptake in adipocytes.J. Cell Sci. 2015; 128: 472-486Crossref PubMed Scopus (65) Google Scholar]. Identifying the exact mechanisms involved in cAMP-induced modulation of SIKs and subsequent biological actions represents a key future challenge.Dentin et al. reported that in hepatocytes, insulin promotes SIK2 phosphorylation on Ser358 and that this results in stimulation of SIK2 activity measured in vitro [24Dentin R. et al.Insulin modulates gluconeogenesis by inhibition of the coactivator TORC2.Nature. 2007; 449: 366-369Crossref PubMed Scopus (324) Google Scholar]. However, these observations have been challenged by more recent studies that found no phosphorylation of Ser358 in response to insulin, and no effect of cAMP-induction or insulin on the in vitro activity of SIK2 [20Henriksson E. et al.The AMPK-related kinase SIK2 is regulated by cAMP via phosphorylation at Ser358 in adipocytes.Biochem. J. 2012; 444: 503-514Crossref PubMed Scopus (49) Google Scholar, 21Patel K. et al.The LKB1-salt-inducible kinase pathway functions as a key gluconeogenic suppressor in the liver.Nat. Commun. 2014; 5: 4535-4550Crossref PubMed Scopus (104) Google Scholar]. The effect of feeding signals on SIK function thus remains controversial (see ‘Liver’). During 1999–2000, Sik was identified as a gene that is rapidly induced in response to various stimuli in different cell types. Upon identification of additional isoforms, this gene and its gene product were later denoted Sik1. For clarification, the first SIK isoform to be identified is termed SIK1 throughout this review article. Wang et al. [4Wang Z. et al.Cloning of a novel kinase (SIK) of the SNF1/AMPK family from high salt diet-treated rat adrenal.FEBS Lett. 1999; 453: 135-139Crossref PubMed Scopus (118) Google Scholar] sought to identify factors that regulate function of the adrenal cortex in response to changes in plasma Na+/K+ balance and screened genes that were induced in rat adrenal gland following a Na+- or K+-enriched diet. A novel cDNA clone, termed salt-inducible kinase (Sik), encoding a 776-amino acid polypeptide with substantial similarity to the AMPK/sucrose non-fermenting kinase (SNF1, the yeast homolog of mammalian AMPK) family, emerged from the screen. They also found that Sik1 mRNA expression is rapidly stimulated by adrenocorticotropic hormone in mouse and rat adrenal cells. Feldman et al. [5Feldman J.D. et al.The salt-inducible kinase, SIK, is induced by depolarization in brain.J. Neurochem. 2000; 74: 2227-2238Crossref PubMed Scopus (47) Google Scholar] were searching for genes in neuronal cells that were preferentially induced by membrane depolarization. They identified a gene product that is induced by potassium, forskolin, or a calcium ionophore and hence termed it KID-2 (kinase-induced by depolarization-2). Xia et al. [6Xia Y. et al.The new serine-threonine kinase, Qik, is a target of the Qin oncogene.Biochem. Biophys. Res. Commun. 2000; 276: 564-570Crossref PubMed Scopus (10) Google Scholar] were studying the role of Qin, a gene shown to induce oncogenic transformation, and sought to identify genes that were differentially expressed in chicken embryo fibroblasts transformed by Qin. One of the up-regulated Qin targets identified was a protein kinase termed QIK (Qin-induced kinase) that displayed high homology to SIK1 cloned by Wang et al. [4Wang Z. et al.Cloning of a novel kinase (SIK) of the SNF1/AMPK family from high salt diet-treated rat adrenal.FEBS Lett. 1999; 453: 135-139Crossref PubMed Scopus (118) Google Scholar] just before their publication. It should also be noted that in 1994, Ruiz and colleagues [7Ruiz J.C. et al.Identification of novel protein kinases expressed in the myocardium of the developing mouse heart.Mech. Dev. 1994; 48: 153-164Crossref PubMed Scopus (48) Google Scholar] had also identified a putative serine/threonine kinase most similar to SNF1, named myocardial SNF-like kinase (msk). Identification of Sik1 prompted Okamoto and colleagues to search for its closely related genes. Analysis of human and mouse genomic databases led them to identify such genes, which they termed Sik2 and Sik3 [8Horike N. et al.Adipose-specific expression, phosphorylation of Ser794 in insulin receptor substrate-1, and activation in diabetic animals of salt-inducible kinase-2.J. Biol. Chem. 2003; 278: 18440-18447Crossref PubMed Scopus (118) Google Scholar, 9Screaton R.A. et al.The CREB coactivator TORC2 functions as a calcium- and cAMP-sensitive coincidence detector.Cell. 2004; 119: 61-74Abstract Full Text Full Text PDF PubMed Scopus (511) Google Scholar]. The Sik1 gene is located on human chromosome 21, whereas both Sik2 and Sik3 genes are localized on chromosome 11. SIKs are evolutionary conserved; Drosophila melanogaster and Caenorhabditis elegans both have SIK homologs, termed dSIK and Kin-29, respectively [13Lanjuin A. Sengupta P. Regulation of chemosensory receptor expression and sensory signaling by the KIN-29 Ser/Thr kinase.Neuron. 2002; 33: 369-381Abstract Full Text Full Text PDF PubMed Scopus (48) Google Scholar, 14Okamoto M. et al.Salt-inducible kinase in steroidogenesis and adipogenesis.Trends Endocrinol. Metab. 2004; 15: 21-26Abstract Full Text Full Text PDF PubMed Scopus (72) Google Scholar, 15Wang B. et al.The insulin-regulated CREB coactivator TORC promotes stress resistance in Drosophila.Cell Metab. 2008; 7: 434-444Abstract Full Text Full Text PDF PubMed Scopus (75) Google Scholar, 16Choi S. et al.Drosophila salt-inducible kinase (SIK) regulates starvation resistance through cAMP-response element-binding protein (CREB)-regulated transcription coactivator (CRTC).J. Biol. Chem. 2011; 286: 2658-2664Crossref PubMed Scopus (39) Google Scholar]. All three SIK isoforms share a highly homologous N-terminal kinase domain, but C-terminal regions are less conserved and differ in length (Figure I). SIK isoforms also possess a ubiquitin-associated domain (Figure I) that appears to play a role in allowing SIKs (and other AMPK-related kinases) to be activated by LKB1 [17Jaleel M. et al.Identification of the sucrose non-fermenting related kinase SNRK, as a novel LKB1 substrate.FEBS Lett. 2005; 579: 1417-1423Crossref PubMed Scopus (116) Google Scholar, 18Rider M.H. The ubiquitin-associated domain of AMPK-related protein kinases allows LKB1-induced phosphorylation and activation.Biochem. J. 2006; 394: e7-e9Crossref PubMed Scopus (6) Google Scholar]. Like other AMPK-related kinases, SIKs are phosphorylated by LKB1 on a highly conserved threonine residue (homologous to Thr172 in AMPK) in their activation loop [1Lizcano J.M. et al.LKB1 is a master kinase that activates 13 kinases of the AMPK subfamily, including MARK/PAR-1.EMBO J. 2004; 23: 833-843Crossref PubMed Scopus (1058) Google Scholar, 17Jaleel M. et al.Identification of the sucrose non-fermenting related kinase SNRK, as a novel LKB1 substrate.FEBS Lett. 2005; 579: 1417-1423Crossref PubMed Scopus (116) Google Scholar]. This phosphorylation is essential for their catalytic activity, but not known to be affected in response to external stimuli or by low cellular energy conditions [1Lizcano J.M. et al.LKB1 is a master kinase that activates 13 kinases of the AMPK subfamily, including MARK/PAR-1.EMBO J. 2004; 23: 833-843Crossref PubMed Scopus (1058) Google Scholar, 19Sakamoto K. et al.Activity of LKB1 and AMPK-related kinases in skeletal muscle: effects of contraction, phenformin, and AICAR.Am. J. Physiol. Endocrinol. Metab. 2004; 287: E310-E317Crossref PubMed Scopus (264) Google Scholar] like AMPK. The most widely received model is that LKB1 is constitutively active; therefore, its dependent kinases (except AMPK) are also constitutively phosphorylated (on the activation loop) and active in cells. Multiple studies have shown that SIKs are phosphorylated at a C-terminal serine residue (SIK1, Ser577; SIK2, Ser587; SIK3, Ser551) in response to agents that elevate intracellular cAMP levels and thus activate protein kinase A (PKA) [10Takemori H. et al.ACTH-induced nucleocytoplasmic translocation of salt-inducible kinase. Implication in the protein kinase A-activated gene transcription in mouse adrenocortical tumor cells.J .Biol. Chem. 2002; 277: 42334-42343Crossref PubMed Scopus (63) Google Scholar, 20Henriksson E. et al.The AMPK-related kinase SIK2 is regulated by cAMP via phosphorylation at Ser358 in adipocytes.Biochem. J. 2012; 444: 503-514Crossref PubMed Scopus (49) Google Scholar, 21Patel K. et al.The LKB1-salt-inducible kinase pathway functions as a key gluconeogenic suppressor in the liver.Nat. Commun. 2014; 5: 4535-4550Crossref PubMed Scopus (104) Google Scholar, 22Sonntag T. et al.14-3-3 proteins mediate inhibitory effects of cAMP on salt-inducible kinases (SIKs).FEBS J. 2018; 285: 467-480Crossref PubMed Scopus (30) Google Scholar, 23Berggreen C. et al.cAMP-elevation mediated by beta-adrenergic stimulation inhibits salt-inducible kinase (SIK) 3 activity in adipocytes.Cell Signal. 2012; 24: 1863-1871Crossref PubMed Scopus (23) Google Scholar]. Detailed studies of glucagon- and catecholamine-induced phosphorylation of SIK2 have been performed in hepatocytes and adipocytes, respectively, and these studies report additional PKA phosphorylation sites (Ser343, Ser358, Thr484, and Ser587) [20Henriksson E. et al.The AMPK-related kinase SIK2 is regulated by cAMP via phosphorylation at Ser358 in adipocytes.Biochem. J. 2012; 444: 503-514Crossref PubMed Scopus (49) Google Scholar, 21Patel K. et al.The LKB1-salt-inducible kinase pathway functions as a key gluconeogenic suppressor in the liver.Nat. Commun. 2014; 5: 4535-4550Crossref PubMed Scopus (104) Google Scholar]. A large body of evidence indicates that cAMP-induced phosphorylation serves to restrict SIK cellular activity [9Screaton R.A. et al.The CREB coactivator TORC2 functions as a calcium- and cAMP-sensitive coincidence detector.Cell. 2004; 119: 61-74Abstract Full Text Full Text PDF PubMed Scopus (511) Google Scholar, 10Takemori H. et al.ACTH-induced nucleocytoplasmic translocation of salt-inducible kinase. Implication in the protein kinase A-activated gene transcription in mouse adrenocortical tumor cells.J .Biol. Chem. 2002; 277: 42334-42343Crossref PubMed Scopus (63) Google Scholar, 24Dentin R. et al.Insulin modulates gluconeogenesis by inhibition of the coactivator TORC2.Nature. 2007; 449: 366-369Crossref PubMed Scopus (324) Google Scholar, 25Muraoka M. et al.Involvement of SIK2/TORC2 signaling cascade in the regulation of insulin-induced PGC-1alpha and UCP-1 gene expression in brown adipocytes.Am. J. Physiol. Endocrinol. Metab. 2009; 296: E1430-E1439Crossref PubMed Scopus (41) Google Scholar], however with mechanisms that differ between SIK isoforms and that in several aspects remain unclear. Rather than a change in intrinsic activity, altered substrate availability is likely involved. Indeed, SIK1 undergoes phosphorylation-dependent nucleocytoplasmic shuttling [10Takemori H. et al.ACTH-induced nucleocytoplasmic translocation of salt-inducible kinase. Implication in the protein kinase A-activated gene transcription in mouse adrenocortical tumor cells.J .Biol. Chem. 2002; 277: 42334-42343Crossref PubMed Scopus (63) Google Scholar]. Moreover, the phosphorylation of SIK2 and SIK3 was shown to induce their binding to 14-3-3 proteins, which coincides with reduced substrate binding [20Henriksson E. et al.The AMPK-related kinase SIK2 is regulated by cAMP via phosphorylation at Ser358 in adipocytes.Biochem. J. 2012; 444: 503-514Crossref PubMed Scopus (49) Google Scholar, 22Sonntag T. et al.14-3-3 proteins mediate inhibitory effects of cAMP on salt-inducible kinases (SIKs).FEBS J. 2018; 285: 467-480Crossref PubMed Scopus (30) Google Scholar, 26Henriksson E. et al.SIK2 regulates CRTCs, HDAC4 and glucose uptake in adipocytes.J. Cell Sci. 2015; 128: 472-486Crossref PubMed Scopus (65) Google Scholar]. Identifying the exact mechanisms involved in cAMP-induced modulation of SIKs and subsequen